CN110672485A - Method for accurately measuring surface hydrophobicity of activated sludge by adsorbing dye - Google Patents
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- 239000010802 sludge Substances 0.000 title claims abstract description 121
- 238000000034 method Methods 0.000 title claims abstract description 42
- 238000001179 sorption measurement Methods 0.000 claims abstract description 22
- 239000007853 buffer solution Substances 0.000 claims abstract description 18
- XJCPMUIIBDVFDM-UHFFFAOYSA-M nile blue A Chemical compound [Cl-].C1=CC=C2C3=NC4=CC=C(N(CC)CC)C=C4[O+]=C3C=C(N)C2=C1 XJCPMUIIBDVFDM-UHFFFAOYSA-M 0.000 claims abstract description 16
- 239000000243 solution Substances 0.000 claims abstract description 16
- 239000012528 membrane Substances 0.000 claims abstract description 13
- 239000000872 buffer Substances 0.000 claims abstract description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 56
- 229910052757 nitrogen Inorganic materials 0.000 claims description 28
- 238000005406 washing Methods 0.000 claims description 27
- 239000000523 sample Substances 0.000 claims description 25
- 239000006228 supernatant Substances 0.000 claims description 21
- 238000005119 centrifugation Methods 0.000 claims description 12
- 239000000725 suspension Substances 0.000 claims description 11
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 7
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 6
- 238000002835 absorbance Methods 0.000 claims description 6
- 239000013595 supernatant sample Substances 0.000 claims description 6
- 239000012153 distilled water Substances 0.000 claims description 5
- 230000002779 inactivation Effects 0.000 claims description 5
- 239000007788 liquid Substances 0.000 claims description 5
- 230000029087 digestion Effects 0.000 claims description 4
- 238000001914 filtration Methods 0.000 claims description 4
- USHAGKDGDHPEEY-UHFFFAOYSA-L potassium persulfate Chemical compound [K+].[K+].[O-]S(=O)(=O)OOS([O-])(=O)=O USHAGKDGDHPEEY-UHFFFAOYSA-L 0.000 claims description 4
- 229910021578 Iron(III) chloride Inorganic materials 0.000 claims description 3
- 239000012535 impurity Substances 0.000 claims description 3
- 238000000926 separation method Methods 0.000 claims description 3
- 238000006243 chemical reaction Methods 0.000 claims description 2
- 238000007865 diluting Methods 0.000 claims description 2
- 238000002156 mixing Methods 0.000 claims description 2
- 239000000047 product Substances 0.000 claims description 2
- 230000000274 adsorptive effect Effects 0.000 claims 1
- 238000004140 cleaning Methods 0.000 abstract description 4
- PXIPVTKHYLBLMZ-UHFFFAOYSA-N sodium azide Substances [Na+].[N-]=[N+]=[N-] PXIPVTKHYLBLMZ-UHFFFAOYSA-N 0.000 abstract description 4
- 238000010561 standard procedure Methods 0.000 abstract 1
- 238000005259 measurement Methods 0.000 description 7
- 210000004027 cell Anatomy 0.000 description 5
- YNPNZTXNASCQKK-UHFFFAOYSA-N phenanthrene Chemical compound C1=CC=C2C3=CC=CC=C3C=CC2=C1 YNPNZTXNASCQKK-UHFFFAOYSA-N 0.000 description 4
- 239000010865 sewage Substances 0.000 description 4
- 230000000875 corresponding effect Effects 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 150000001335 aliphatic alkanes Chemical class 0.000 description 2
- 230000015556 catabolic process Effects 0.000 description 2
- 238000006731 degradation reaction Methods 0.000 description 2
- 239000007888 film coating Substances 0.000 description 2
- 238000009501 film coating Methods 0.000 description 2
- 125000000524 functional group Chemical group 0.000 description 2
- 238000000691 measurement method Methods 0.000 description 2
- 238000002798 spectrophotometry method Methods 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 1
- 238000004566 IR spectroscopy Methods 0.000 description 1
- 208000012868 Overgrowth Diseases 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000006065 biodegradation reaction Methods 0.000 description 1
- 230000032770 biofilm formation Effects 0.000 description 1
- 230000031018 biological processes and functions Effects 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 230000018044 dehydration Effects 0.000 description 1
- 238000006297 dehydration reaction Methods 0.000 description 1
- 238000005189 flocculation Methods 0.000 description 1
- 230000016615 flocculation Effects 0.000 description 1
- 238000005187 foaming Methods 0.000 description 1
- 125000001165 hydrophobic group Chemical group 0.000 description 1
- 230000005661 hydrophobic surface Effects 0.000 description 1
- 230000000415 inactivating effect Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000004062 sedimentation Methods 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 239000002351 wastewater Substances 0.000 description 1
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- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume or surface-area of porous materials
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- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
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- G01N21/33—Investigating relative effect of material at wavelengths characteristic of specific elements or molecules, e.g. atomic absorption spectrometry using ultraviolet light
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Abstract
The invention discloses a method for accurately measuring the surface hydrophobicity of activated sludge by using an adsorption dye. The method comprises the steps of centrifuging collected sludge samples for three times by using PBS buffer solution, and centrifuging the sludge samples by using NaN3Inactivating; then NaN was dissolved in PBS buffer3Cleaning; suspending with a nile blue solution; respectively taking sludge samples which are suspended for 0min and suspended for a period of time, centrifuging the sludge samples in a certain volume, and passing supernate of the centrifuged samples through a membrane; measuring the TN content of the solution after the membrane is coated by adopting a national standard method; finally, a phase with activated sludge surface hydrophobicity equal to the concentration of TN of the two supernatantsFor the variance. The method can accurately measure the surface hydrophobicity of the activated sludge, and is an improved method for measuring the surface hydrophobicity of the activated sludge.
Description
Technical Field
The invention relates to a method for accurately measuring the surface hydrophobicity of activated sludge by adsorbing dye, belonging to the field of biological treatment of wastewater.
Background
Cell surface hydrophobicity is one of the important physicochemical parameters that influence cell surface interactions. In the sewage treatment process, the hydrophobicity of cells has an important influence on the flocculation, sedimentation and dehydration processes of activated sludge. Thus, the change in the hydrophobicity of cells in the activated sludge may cause a serious operational problem in the sewage treatment plant. For example, the sludge foaming process is mainly due to overgrowth of filamentous bacteria with extremely hydrophobic surfaces. Furthermore, for attachment-growth biological processes, such as trickling, cell hydrophobicity can also affect biofilm formation to a large extent. Therefore, in order to optimize the biological treatment process of sewage in sewage treatment plants, it is necessary to precisely measure the hydrophobicity of activated sludge.
The current methods for representing hydrophobicity mainly comprise three methods, namely a contact angle measurement method, an alkane distribution method and a phenanthrene adsorption method. Wherein, the contact angle measurement can only measure the contact angle in a certain range on the surface of the sludge, and has certain limitation; when the alkane distribution method is used for measuring, the sludge is crushed, and the hydrophobicity of the surface of the sludge cannot be measured; the specific operation of the phenanthrene adsorption method is not yet clear. Therefore, we developed a method for accurately determining the surface hydrophobicity of activated sludge by adsorption of dye by sludge.
Disclosure of Invention
The invention aims to provide a method for accurately measuring the surface hydrophobicity of activated sludge by adsorbing dye.
The method for accurately measuring the surface hydrophobicity of the activated sludge by adsorbing the dye mainly comprises the following steps of (1) collecting and cleaning a sludge sample; (2) sludge inactivation: the microbial degradation is eliminated, and only the adsorption of the nitrogen in the dye by the sludge surface is considered; (3) washing sludge; (4) suspending the dye: adding a Nile blue solution into a sludge sample, wherein the solvent of the Nile blue is PBS; suspending for 0min, centrifuging the sample; suspending for 30min, sampling again, and centrifuging; (5) film coating: passing the centrifuged sample supernatant through a water film to ensure the accuracy of subsequent total nitrogen measurement; (6) TN measurement; (7) the hydrophobicity of the activated sludge surface was calculated.
The technical scheme adopted by the invention is as follows:
a method for accurately measuring the surface hydrophobicity of activated sludge by using an adsorption dye comprises the following steps:
step 1: quantitatively transferring a sludge sample into a centrifugal tube, and performing centrifugal solid-liquid separation; then, continuously washing and centrifugally separating the separated sludge for multiple times by using PBS buffer solution;
step 2: quantitatively transferring PBS buffer solution, completely suspending the centrifugally separated sludge, and adding NaN into the suspension3Placing the sludge in a shaking table and shaking for a certain time to inactivate the sludge;
and step 3: quantitatively transferring the inactivated sludge sample, repeatedly washing and centrifuging for several times by using PBS buffer solution to remove NaN in the sludge3Until the washing solution and FeCl3The solution does not generate red complex after mixing;
and 4, step 4: quantitatively transferring the nile blue solution, and suspending the sludge washed and centrifuged in the step 3; immediately taking part of suspended sludge for centrifugation when suspension is finished for 0min, and reserving supernatant for later use; oscillating the residual suspended sludge in a shaking table and suspending for 30min, centrifuging part of the suspended sludge, and keeping the supernatant for later use;
and 5: respectively filtering the supernatant after suspending for 0min and 30min through 0.45 micron filter membranes;
step 6: respectively measuring the total nitrogen concentration of the two supernatants obtained after the filtration in the step 5;
and 7: and calculating the relative change rate of the total nitrogen concentration of the supernatant after suspending for 0min and 30min, and using the relative change rate to represent the surface hydrophobicity of the activated sludge.
Based on the scheme, the following preferred schemes can be adopted for parameters and materials in each step:
preferably, the specific method in step 1 is as follows: taking 30mL of sludge sample in a centrifuge tube, centrifuging for 5min at 10000rpm, and removing impurities in the sludge sample; after which washing and centrifugation were repeated three more times with PBS buffer.
Preferably, in step 1, the amount of the PBS buffer solution used in each washing and centrifuging process is 10mL, the centrifugation speed is 10000rpm, and the centrifugation time is 5 min.
Preferably, in the step 1, the sludge concentration of the sludge sample is 3000-6000 mg/L.
Preferably, in step 2, the PBS buffer solution is quantitatively removed in a volume of 100mL, and NaN is added to the suspension3The volume was 1 mL.
Preferably, in the step 2, the reaction conditions are 200rpm, 25 ℃ and 30min when the inactivation is carried out in a shaking table.
Preferably, in the step 3, 30mL of inactivated sludge sample is taken, washing and centrifugation are repeatedly carried out by using PBS buffer solution, and 1-2 drops of 100g/L FeCl are added into the last washing liquid3If the red complex does not appear, NaN in the sludge is indicated3And washing the product.
Preferably, in the step 4, the concentration of the nile blue solution used for suspension is 20mg/L, and the volume is 60 mL.
Preferably, the membrane in the step 5 adopts a needle filter and a 0.45 micron filter membrane.
Preferably, in step 6, the method for measuring total nitrogen is as follows: taking 5mL of supernatant sample after the membrane is filtered by a pipette, and adding the supernatant sample into a 25mL ground colorimetric tube; adding distilled water to 10 mL; adding 5mL of alkaline potassium persulfate, binding the pipe orifice with gauze, and digesting; after digestion, 1mL of 1+9 hydrochloric acid is added, and the volume is adjusted to 25m by using distilled waterL, measuring the absorbance A at wavelengths of 220nm and 275nm respectively220、A275According to the corrected absorbance A ═ A220-2A275And calculating to obtain the corresponding total nitrogen concentration by combining the standard curve.
The invention has the beneficial effects that:
according to the method, on the basis of the traditional method for measuring the surface hydrophobicity of the activated sludge, the relative change quantity of the total nitrogen concentration before and after adsorption of the dye (Nile blue) is measured through the adsorption effect of the activated sludge on the dye, and the surface hydrophobicity of the activated sludge is accurately measured, so that the follow-up effective research is facilitated.
Drawings
FIG. 1 is a flow chart for accurately measuring the surface hydrophobicity of activated sludge by adsorbing dye.
FIG. 2 shows the addition of 1% NaN to a sludge sample30min and 30min infrared spectrogram.
Detailed Description
The invention is further illustrated by the following figures and specific examples.
As shown in fig. 1, a method for accurately measuring the surface hydrophobicity of activated sludge by using an adsorption dye generally comprises the following basic steps: (1) collecting and cleaning a sludge sample; (2) inactivating the sludge; (3) washing sludge; (4) suspending the dye; (5) film coating; (6) TN measurement; (7) the hydrophobicity of the activated sludge surface was calculated.
The principle of hydrophobicity measurement according to the present invention is as follows:
the activated sludge is a floc of a porous structure and extracellular polymers, has both hydrophobic groups and hydrophilic groups, and has strong adsorption capacity on hydrophilic dye Nile blue. Nile blue is dissolved in PBS buffer solution, and the content of nitrogen in the 20mg/L Nile blue PBS buffer solution is 2.292 mg/L. In order to eliminate the biodegradation effect of sludge on nitrogen in Nile blue, NaN is used before dye suspension3Inactivation, NaN3Has no influence on the functional groups on the surface of the sludge. The hydrophobicity of the surface of the sludge is positively correlated with the adsorption capacity of the sludge to Nile blue, and the hydrophobicity of the surface of the sludge is expressed by measuring the relative change amount of the nitrogen content in the aqueous solution before and after the adsorption of the sludge. The total nitrogen can adopt national standard (HJ 636-2012) basic oversulfidePotassium is resolved and is measured by an ultraviolet spectrophotometry.
The specific implementation of each step in the method is described in detail below:
(1) step 1, sludge sample collection and cleaning
30mL of sludge sample (the sludge concentration is 6000mg/L) is measured by a measuring cylinder and put into a 50mL centrifuge tube, the centrifuge tube is centrifuged at 10000rpm for 5min, the suspended impurities in the sludge sample are removed, and the supernatant is discarded. Then, 10mL of PBS buffer solution is measured by a pipette gun and mixed with the precipitated sludge after the previous centrifugation, the mixture is centrifuged again at 10000rpm for 5min, and the mixing-centrifugation process is repeated for 3 times, so that a good environment is provided for ensuring the structural and functional integrity of the sludge for the next suspension.
(2) Step 2, sludge inactivation
Suspending the centrifuged sludge sample into a 250mL conical flask by using 100mL of PBS buffer solution, and adding 1mL of NaN3Then, the inactivation of the sludge is realized in a shaking table at 200rpm and 25 ℃ for 30 min. The addition of NaN was found by IR spectroscopy according to FIG. 23Last 0min and 30min, NaN3Has little influence on the functional groups on the surface of the sludge, so NaN is adopted3Inactivation can eliminate the degradation effect of microorganisms on nitrogen in the dye under the condition of not damaging the surface structure of the sludge, and only considers the adsorption of the sludge surface on the nitrogen in the dye.
(3) Step 3, sludge washing
Taking 30mL of inactivated sludge, washing with PBS buffer solution for three times, and adding NaN in the sludge3Washing off to prevent influence on subsequent tests, centrifuging after each washing, and performing solid-liquid separation. Adding 1-2 drops of 100g/L FeCl into the washing solution separated after the last washing3If the washing solution is mixed with FeCl3The solution does not generate red complex after being mixed, which indicates that NaN in the sludge3The washing is completed, otherwise, the washing and the centrifugation are required to be repeated again.
(4) Step 4, suspending the dye
60mL of nile blue solution with the concentration of 20mg/L is measured by a measuring cylinder, and the sludge after the previous step of centrifugation is completely suspended in a 150mL conical flask. When the suspension is finished for 0min (namely when the suspension is just finished), 10mL of suspended sludge sample is immediately taken to be centrifuged at 10000rpm for 5min, and the supernatant is reserved for later use.
And (3) oscillating the residual suspended sludge in a shaking table and continuing to suspend for 30min, centrifuging 10mL of suspended sludge sample at 10000rpm for 5min, and keeping the supernatant for later use. Wherein, the prepared 1L of Nile blue solution contains 2.292mg of total nitrogen, and the measurement range of the total nitrogen in the national standard is 0.050-4 mg/L, so the hydrophobicity of the sludge surface is shown by measuring the relative change of the concentration of the total nitrogen in Nile blue adsorbed on the sludge surface for 0min and 10 min.
(5) Step 5, coating the membrane
And (3) taking 10mL of the centrifuged two supernatants (respectively suspending for 0min and 30min), and passing through a 0.45-micrometer water system filter membrane by using a needle filter to remove suspended sludge in the supernatants so as to ensure the accuracy of the total nitrogen determination.
(6) Step 6, TN measurement
For the two filtered supernatants, total nitrogen is measured by an alkaline potassium persulfate digestion ultraviolet spectrophotometry method, which comprises the following specific steps:
taking 5mL of supernatant sample after the membrane is filtered by a pipette, and adding the supernatant sample into a 25mL ground colorimetric tube; adding distilled water to 10 mL; adding 5mL of alkaline potassium persulfate, binding the pipe orifice with gauze, and digesting; after digestion, adding 1mL of 1+9 hydrochloric acid, diluting to 25mL with distilled water, and measuring absorbance A at wavelength of 220nm and 275nm respectively220、A275According to the corrected absorbance A ═ A220-2A275And calculating to obtain the corresponding total nitrogen concentration by combining the standard curve.
(7) Step 7, calculating the hydrophobicity of the surface of the activated sludge
And calculating the relative change rate of the total nitrogen concentration of the supernatant after suspending for 0min and 30min, and using the relative change rate to represent the surface hydrophobicity of the activated sludge. The surface hydrophobicity of the activated sludge is (total nitrogen concentration in supernatant suspended for 0 min-total nitrogen concentration in supernatant suspended for 30 min)/total nitrogen concentration in supernatant suspended for 0 min.
This example sets up A, B, C three sets of parallel tests, and the final calculated surface hydrophobicity results are as follows: relative change rate of total nitrogen concentration of activated sludge surface hydrophobicity
The variance of the three sets of data is 0.006668, indicating that the measurement method has better accuracy and reproducibility. It should be noted that the sludge surface hydrophobicity refers to relative hydrophobicity, different samples can respectively calculate corresponding hydrophobicity values according to the method, and the relative size of the hydrophobicity of the sludge sample is determined by comparison.
Claims (10)
1. The method for accurately measuring the surface hydrophobicity of the activated sludge by using the adsorption dye is characterized by comprising the following steps of:
step 1: quantitatively transferring a sludge sample into a centrifugal tube, and performing centrifugal solid-liquid separation; then, continuously washing and centrifugally separating the separated sludge for multiple times by using PBS buffer solution;
step 2: quantitatively transferring PBS buffer solution, completely suspending the centrifugally separated sludge, and adding NaN into the suspension3Placing the sludge in a shaking table and shaking for a certain time to inactivate the sludge;
and step 3: quantitatively transferring the inactivated sludge sample, repeatedly washing and centrifuging for several times by using PBS buffer solution to remove NaN in the sludge3Until the washing solution and FeCl3The solution does not generate red complex after mixing;
and 4, step 4: quantitatively transferring the nile blue solution, and suspending the sludge washed and centrifuged in the step 3; immediately taking part of suspended sludge for centrifugation when suspension is finished for 0min, and reserving supernatant for later use; oscillating the residual suspended sludge in a shaking table and suspending for 30min, centrifuging part of the suspended sludge, and keeping the supernatant for later use;
and 5: respectively filtering the supernatant after suspending for 0min and 30min through 0.45 micron filter membranes;
step 6: respectively measuring the total nitrogen concentration of the two supernatants obtained after the filtration in the step 5;
and 7: and calculating the relative change rate of the total nitrogen concentration of the supernatant after suspending for 0min and 30min, and using the relative change rate to represent the surface hydrophobicity of the activated sludge.
2. The method for accurately measuring the surface hydrophobicity of the activated sludge by using the adsorption dye according to claim 1, wherein the specific method in the step 1 is as follows: taking 30mL of sludge sample in a centrifuge tube, centrifuging for 5min at 10000rpm, and removing impurities in the sludge sample; after which washing and centrifugation were repeated three more times with PBS buffer.
3. The method for accurately measuring the surface hydrophobicity of the activated sludge by using the adsorption dye as claimed in claim 1, wherein in the step 1, the amount of the PBS buffer solution used in each washing and centrifuging process is 10mL, the centrifuging speed is 10000rpm, and the centrifuging time is 5 min.
4. The method for accurately measuring the surface hydrophobicity of the activated sludge by the adsorptive dye according to claim 1, wherein the sludge concentration of the sludge sample in the step 1 is 3000-6000 mg/L.
5. The method for accurately measuring the surface hydrophobicity of the activated sludge by using the adsorption dye as claimed in claim 1, wherein in the step 2, the volume of the PBS buffer solution is quantitatively removed to be 100mL, and NaN is added into the suspension3The volume was 1 mL.
6. The method for accurately measuring the surface hydrophobicity of the activated sludge by using the adsorption dye according to claim 1, wherein the reaction conditions in the inactivation in the shaking table in the step 2 are 200rpm, 25 ℃ and 30 min.
7. The method for accurately measuring the surface hydrophobicity of the activated sludge by using the adsorption dye as claimed in claim 1, wherein in the step 3, 30mL of the inactivated sludge sample is taken, the washing and the centrifugation are repeatedly carried out by using the PBS buffer solution, and 1-2 drops of 100g/L FeCl are added into the last washing liquid3If the red complex does not appear, NaN in the sludge is indicated3And washing the product.
8. The method for accurately measuring the surface hydrophobicity of the activated sludge by the adsorption dye according to claim 1, wherein the nile blue solution used for suspending in the step 4 has a concentration of 20mg/L and a volume of 60 mL.
9. The method for accurately measuring the surface hydrophobicity of the activated sludge by using the adsorption dye as claimed in claim 1, wherein the membrane in the step 5 adopts a needle filter and a 0.45 micron filter membrane.
10. The method for accurately measuring the surface hydrophobicity of the activated sludge by the adsorption dye according to claim 1, wherein in the step 6, the total nitrogen is measured by the following method: taking 5mL of supernatant sample after the membrane is filtered by a pipette, and adding the supernatant sample into a 25mL ground colorimetric tube; adding distilled water to 10 mL; adding 5mL of alkaline potassium persulfate, binding the pipe orifice with gauze, and digesting; after digestion, adding 1mL of 1+9 hydrochloric acid, diluting to 25mL with distilled water, and measuring absorbance A at wavelength of 220nm and 275nm respectively220、A275According to the corrected absorbance A ═ A220-2A275And calculating to obtain the corresponding total nitrogen concentration by combining the standard curve.
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